Vaccination of European Sea Bass (Dicentrarchus labrax) against Viral Nervous Necrosis and Characterization of Protective Immunity

European sea bass (Dicentrarchus labrax) is one of the main cultured fish species in the Mediterranean with an increasing production every year. Unfortunately, sea bass are susceptible to the disease viral nervous necrosis (VNN) which can cause great losses compromising both fish welfare and the economy of the production. Nervous necrosis virus (NNV) is the etiologic agent. It is a small single stranded bi-segmented RNA virus belonging to the genus Betanodavirus. It infects many marine and fresh water species and thus circulate both in farmed and wild fish, making prevention and elimination difficult. Vaccination, breeding towards resistant lines of fish and biosecurity measures seems to be the most promising management tools available to the farmer. This thesis investigates the first two through vaccination and challenge experiments of European sea bass. The thesis includes three manuscripts and two unpublished studies.

Natural resistance to the disease was investigated in a challenge experiment where three distinct populations of European sea bass were infected with two different variants of NNV. These results are presented in manuscript I. We found that the population originating from the Eastern Mediterranean had a significant higher survival to challenge with a pathogenic redspotted grouper (RG)-NNV strain. This strain is circulating in the eastern Mediterranean, and the resistance therefore might be because of a co-evolutionary advantage of this population. The mechanisms seems to be related to an ability to limit the entry into and/or replication of virus.

Vaccine-induced protection against viral nervous necrosis in European sea bass was investigated through experimental testing of a virus-like particle (VLP)-based vaccine in vaccination-challenge experiments. This work is presented in manuscripts II and III and two unpublished studies. An RGNNV VLP was made available through a partner, W42 biotechnology, and the efficacy of different doses and formulations were tested. The vaccine induced immune-response was evaluated through serology and immune gene expression analysis. Protection against disease was evaluated in experimental challenge. Furthermore, focus was on testing the duration of immunity, since this aspect is important for the applied potential of the vaccine.

The VLP-based vaccine induced a dose-dependent and long lasting response of neutralizing antibodies. Furthermore, VLP-vaccinated fish down to a size of 5 g was significantly protected in experimental intramuscular challenge with RGNNV up to 7.5 months after vaccination. The average level of neutralizing antibodies correlate with protection of the given group, although a high neutralizing titer alone was not enough to protect the individua fish. Both the innate and adaptive immune systems were activated, which might explain the high efficacy that could be obtained, even without an adjuvant. The VLP-vaccinated fish had a better ability to control the infection, as a lower prevalence of virus was detected in the brain of VLP-vaccinated survivors compared to mock-vaccinated survivors. A small pilot study with oral delivery of the VLP was unsuccessful, although the potential of this delivery route cannot be fully ruled out based on this small pilot study.

In conclusion, the NNV VLP-based vaccine harbour great potential in the prevention of VNN in sea bass. Future studies could be aimed at investigating alternative delivery routes and testing the already effective intraperitoneal delivery route in a field setting. The protective effect against other strains of RGNNV and in other NNV susceptible fish species could also be investigated, which would, if successful, increase the potential of the vaccine.